This is a new R01 grant application responsive to PA-99-067 "HIV therapeutics: targeting research gaps (3) defining and validating the role of innate immune responses to HIV". The goal of this project is to determine the effects of HIV infection on the natural killer T (NKT) cell compartment. NKT cells represent a rare subset of lymphocytes defined by uniform expression of T cell receptor alpha-chain segment 24 (Valpha24) and the natural killer cell marker CD161. These cells recognize glycolipids presented by CDld, and have important immunoregulatory functions. Our preliminary studies indicate that the Valpha24 NKT cell compartment is skewed in patients infected with HIV. Our preliminary data indicate that CD4+ and CD4- sub-populations of NKT cells can be identified in peripheral blood and that these cells respond to CDld-presented antigens by producing interferon (IFN)-gamma ex vivo. In our first specific aim we will investigate the expression of surface markers linked to homing, effector function, and memory in NKT cells by flow cytometry, with special focus on differences between CD4+ and CD4- subsets, and assess the function of these cells using an ex vivo cytokine flow cytometric assay. We will also determine the effect of HIV infection on the phenotype and function of the Valpha24 NKT cell compartment and the sensitivity of NKT cells to infection with R5 and X4 viruses. In the second specific aim, we will investigate the effect of HIV on the Valpha24 NKT cell compartment in patients with acute and chronic HIV-1 infection. We will further investigate the relationship between NKT cell activity and opportunistic infections. In the third specific aim, we will ascertain the effect of HIV infection of the thymus on Valpha24 NKT cell development in the SC1D-hu Thy/Liv mouse model. We will study SCID-hu Thy/Liv mice infected with X4 and R5 HIV strains, and determine the impact on NKT cell development. Furthermore, we will assess the presence and function of Valpha24 NKT cells in the periphery of SCID-hu Thy/Liv mice, and assess the response to the CD 1d-presented NKT cell ligand alphaGalCer in vivo. We believe that the proposed studies will contribute considerably to our understanding of NKT cell function in HIV disease, how the virus affects them, and how they may contribute to innate defense against HIV.